Implants comprising biologically derived scaffolds have become important options for tissue/organ repair and regeneration in the treatment of various different diseases and conditions. A continuing and major hurdle is the need to remove antigen-presenting cellular material from the tissue, which tissue then becomes the scaffold. In particular when relatively dense interstitial, connective or supporting tissue, including cartilaginous tissue, such as tracheal tissue, is decellularised, it is very difficult, if not impossible, to remove substantially all antigen-presenting cells from the tissue.
Interstitial, connective and supporting tissue damage, including long tracheal lesions, still represent a challenge to the surgeon. For example, damage resulting from congenital defects, trauma or tumour that compromise more than 4.5-6 cm or more than 30% of the total tracheal length in children is not treatable via primary closure. Thus these patients are rarely considered as surgically curable and the use of implants derived from tracheal tissue is a desired alternative. Reconstruction with a tracheal conduit would extend surgical indications and improve quality of life.
Recent advances in the field of regenerative medicine hold significant promise especially, with regards to tissue engineered replacement tracheal scaffolds. The ideal replacement should be as close to the natural structure as possible providing stability and non-immunogenic characteristics. To fulfil these criteria significant research resources are being directed towards using biological material as a starting point. Preparing a scaffold for regenerative purpose using either allogenic or xenogenic material requires the complete removal of all antigenicity whilst preserving the extracellular matrix to an extent that it is able to support cell attachment and to provide sufficient rigidity for air ventilation.
Known decellularisation techniques use different chemical and biological reagents to wash out antigen presenting cells and cell particles. One established protocol to decellularise tracheal tissue is based on a detergent-enzymatic-method, in which cells are removed from the tracheal tissue by perfusion with various detergents, enzymes and other reagents. Whilst providing a suitable scaffold that had been successfully transplanted in a handful of cases on compassionate grounds, it has not yet reached standard clinical practice. One reason is the lengthy preparation of the scaffold which takes approximately 3 weeks, and the accompanying risk to the patient caused by the time delay.
A standardized “off-the-shelf” scaffold for clinical use requires not only the correct anatomical, functional and biomechanical characteristics but also the feasibility to be prepared in a suitable time frame. To improve decellularisation of tissues, different methods are available, encompassing different combinations of enzymes and detergents. Since the majority of these reagents are known to alter the extracellular matrix a different approach to known techniques is still desired in order to mitigate or prevent alteration of the extracellular matrix.
As interstitial, connective and supporting tissue, such as cartilage, is a specifically dense tissue, a method is required to deliver the decellularising agents deep into the tissue over a relatively short time and with no impact on the tissue ultrastructure of extra cellular matrix, especially when preparing tracheal implants.
The present invention therefore provides an improved method for producing an implant for tissue/organ repair, especially based on interstitial, connective or supporting tissue, including cartilaginous tissue such as the trachea, in which the implant can be prepared in a relatively short period of time, whilst maintaining a substantially intact extra-cellular matrix with removal of substantially all antigen-presenting cells.